The broad goal of this project is to understand the mechanism of liver cancer stem cell survival and resistance to chemotherapy. Accumulating evidence indicates that liver cancer stem cells (CSC) are the origin of a subset of hepatocellular carcinomas (HCC). HCC patients with a stem cell based tumor have a significantly worse prognosis compared to patients with more differentiated HCC. The central hypothesis is that deregulated growth factor signaling allows stem cells to escape normal apoptotic mechanisms and become CSCs. We have identified liver CSC phenotypes in two pre-tumor chronic liver injury models using CD133 expression. These CD133+ CSCs are resistant to Transforming Growth Factor-( (TGF-()- induced apoptosis. To date, no one has reported an unbiased analysis to identify the strongest liver cancer stem cell immuno-phenotype identified from the many published markers. By identifying the most reliable immuno-phenotype of liver CSC, we can further dissect the factors that influence the loss of cell cycle control and resistance to apoptosis critical to liver cancer progression and chemotherapy resistance. The rationale for the proposed research is that new therapeutic approaches for liver cancer can be developed to target liver CSCs, which maintain the larger neoplasia and account for a significant proportion of chemotherapy failures. We will utilize multiple human hepatocellular carcinoma and cholangiocarcinoma cell lines to investigate the single best cancer stem cell marker linked to deregulation of critical pathways known to influence liver carcinoma progression. Aim 1 Determine the liver cancer stem cell surface marker highly correlated with resistance to chemotherapy and to biologically induced apoptosis. Hypothesis: Resistance to signal mediated apoptosis and chemotherapy agents provides CSCs a mechanism of survival. Aim 2 Test regulatory pathways within specific cancer stem cell populations. Hypothesis: Deregulation of key pathways known to influence HCC progression (RAS/MAPK, PI3K/AKT, (-catenin) will provide a mechanism of survival and apoptosis resistance to liver CSCs. The proposed work is innovative because we propose an unbiased analysis of CSC surface markers coupled with an investigation of mechanisms of survival. The work proposed in Aim 1 will provide us with a more complete understanding of the best liver CSC surface marker. Furthermore, we anticipate that Aim 2 will allow us to understand how specific deregulation of key signaling pathways provides liver CSCs with a survival mechanism. PUBLIC HEALTH RELEVANCE: The broad goal of this project is to understand the mechanism of liver cancer stem cell survival and resistance to chemotherapy. We seek to understand the relationship between liver stem cells and cancer stem cells during chronic liver injury and tumorigenesis. If successful, the potential impact of this award will be realized in the area of liver cancer therapy development.